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Purushottam Jha, Ruslana G. Tytarenko, Himanshu Banda, Bharati Matta, Puran S. Bora, Nalini S. Bora; Complement Mediated Apoptosis Leads to the Loss of Retinal Ganglion Cells in Animal Model of Glaucoma. Invest. Ophthalmol. Vis. Sci. 2011;52(14):516.
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This study investigated the role of complement in the protection of retinal ganglion cells (RGCs) in chronic ocular hypertension model of glaucoma.
Intraocular pressure (IOP) was elevated in the left eye of Lewis rats by laser photocoagulation (two treatments, 7 days apart) of episcleral and limbal veins. Right eye did not receive laser treatment and served as control. Animals were injected with cobra venom factor (CVF) every fifth day starting day 7 after first laser, to deplete the complement system. Control animals received similar treatment with PBS. Animals were sacrificed at six week post-laser. The retina was harvested from laser-treated eyes as well as untreated control eyes and processed for flow cytometry, paraffin embedding, and protein extraction. Paraffin sections were used for immunostaining for glial fibrillary acidic protein (GFAP), Brn3a, complement component C3, complement membrane attack complex (MAC), TUNEL, caspase-8, caspase-9, calcium green, BID and BAD. Western blot analysis was used to confirm the levels of C3, MAC, caspase-8 and caspase-9 in the retina of CVF and PBS-treated rats with elevated IOP.
Levels of C3 split products and MAC were elevated in the retina of eyes with increased IOP and complement depletion reduced the loss of RGCs accompanied by decreased expression of GFAP and reduced MAC deposition. In complement depleted rats with increased IOP, reduced TUNEL, active caspase-8 and active caspase-9 staining was observed in ganglion cell layer and the number of apoptotic cells drastically decreased in retina of these animals. Interestingly, complement depletion also resulted in reduction of calcium influx in the retinal cells of the eyes with increased IOP. Furthermore levels of both BID and BAD in RGCs decreased in the CVF-treated animals with increased IOP.
Together, our results provide evidence that complement mediated apoptosis plays a pivotal role in the loss of RGCs in chronic ocular hypertension model of glaucoma.
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